(W0930-01-03) Developing a Platform for Formulation Screening of High Concentration Monoclonal Antibodies (mAbs)

Purpose: High concentration mAb formulations required for high dose subcutaneous (SC) administration can suffer from high viscosity and protein aggregation. To develop an optimized high concentration mAb formulation, screening of excipients/pH/buffer is often required. However, this screening can be difficult as protein solutions without excipients at concentrations above 200 mg/ml can reach gelling point. We propose to use lyophilization for screening formulations of high concentration antibodies as lyophilized mAb can be reconstituted to the desired protein concentration with any excipient/buffer combination without using a high concentration solution with no excipients.

Methods: The antibody (mAb A) was dialyzed into water at moderate concentration (~100 mg/ml) and pre-lyophilization buffer was added. The resulting solution was lyophilized in 6R vials (1.5 ml in each vial) and reconstituted with the desired screening buffer to required protein concentration (e.g., ~200 mg/ml). We optimized pre-lyophilization buffer and lyophilization method to minimize reconstitution time and high molecular weight (HMW) species. This method was used to screen different reconstitution buffers to minimize solution viscosity while maintaining other parameters described above in acceptable range.

Results: We found that lyophilization of mAb A in water resulted in high level of HMW species. Adding buffer and sucrose (5 mM histidine, 1% sucrose, pH 5.9) or 1% sucrose alone decreased HMW species to acceptable level. Lyophilization cycle was initially optimized by removing secondary drying that was shown to be unnecessary. To decrease the reconstitution time, we tried controlled nucleation and annealing. We found that controlled nucleation at -5 C followed by vacuum stoppering resulted in the shortest reconstitution time and acceptable HMW levels. Annealing was less effective in decreasing reconstitution time. This protocol was implemented to screen different viscosity reducers and it was found that several salts and charged amino acids at 150 mM concentration reduced viscosity of reconstituted solution (220 mg/ml) to acceptable level ( < 30 cps) while preserving short reconstitution time and acceptable levels of HMW species.

Conclusion: A lyophilization-based platform for high concentration mAb formulation screening has been developed and can be implemented to screen formulation excipients (such as, viscosity reducers) and pH/buffers.

Acknowledgements: Conflict of interest: LB, EC, CA, MS and XT are employees of Regeneron Pharmaceuticals.

Figure 1. Optimization of the lyophilization cycle.

Figure 2. Screening of reconstitution buffers (10 mM histidine, 2% sucrose, pH 5.9, 150 mM excipient) using the lyophilization method developed in this study.